Characterization of epidermal stem cells: biology, carcinogenesis, and function

长期以来，实验性皮肤癌的小鼠模型一直认为，表皮干细胞是致癌物的主要靶点，形成潜在的肿瘤群体，最终扩展为肿瘤。我们使用CD34基因敲除(CD34KO)小鼠提供了这一点的实验证据，该实验基于CD34在毛囊干细胞上唯一表达的假设，在介导干细胞对致癌物的反应中发挥动态作用。CD34KO小鼠在皮肤肿瘤的发展中表现出显着的损害，我们将这种表型与肿瘤启动子异常诱导的毛囊周期激活和干细胞增殖联系在一起。此外，根据细胞周期分析和克隆培养中的生长情况，我们有证据表明，缺乏CD34的毛囊干细胞的增殖能力降低，这表明CD34对干细胞的反应性至关重要，同时也强调了毛囊干细胞对肿瘤发展的贡献。我们的实验室是第一个证明CD34在小鼠毛囊干细胞上表达的，通过这一点，我们开发了促进活干细胞浓缩的方法学。早些时候，我们从CD34+角质形成细胞的基因表达分析中提供了证据，在TPA处理小鼠皮肤后，CD34+角质形成细胞中缺失的分裂手/分裂脚1基因(Dss 1)的表达上调，随后已被证明在细胞中具有生长调节作用。我们已经确定Dss1通过直接与蛋白酶体组装的19S调节颗粒的Rpn3/S3亚单位结合来介导其生长调控作用，在蛋白酶体与泛素化底物结合和随后的蛋白质降解中具有重要意义。在这方面，我们首次提供了生化证据表明，HsDSS1的R3IM基序与19S RP/20S CP(核心颗粒)复合体一起，通过与Rpn3/S3的相互作用调节蛋白酶体的组装和功能，并利用多泛素化的p53的一个特定子集作为底物。

The mouse model of experimental skin carcinogenesis has long proposed that epidermal stem cells are a major target of carcinogens and form the latent neoplastic population that ultimately expands into tumors. We have provided experimental evidence of this using the CD34 knockout (CD34KO) mouse, based on the hypothesis that CD34, which is uniquely expressed on hair follicle stem cells, plays a dynamic role in mediating stem cell response to carcinogens. CD34KO mice exhibit a striking impairment in skin tumor development and we have linked this phenotype to aberrant tumor promoter-induced activation of hair follicle cycling and stem cell proliferation. Further, we have evidence suggesting that hair follicle stem cells lacking CD34 have a reduced proliferative potential, based upon cell cycle analysis and growth in clonogenic culture, demonstrating that CD34 is critical for stem cell responsiveness as well underscoring the contribution of hair follicle stem cell contribution to tumor development. Our laboratory was the first to demonstrate that CD34 is expressed on mouse hair follicle stem cells, and through this, we developed methodology that facilitated enrichment for living stem cells. Earlier, we provided evidence from gene expression analysis of CD34+ keratinocytes of upregulation in the expression of the Deleted in Split Hand/Split Foot 1 gene (Dss1) following TPA treatment of mouse skin, which has subsequently been shown to have a growth regulatory role in the cell. We have determined that Dss1 mediates its growth regulatory effects through direct binding with the Rpn3/S3 subunit of the 19S regulatory particle of the proteasome assembly, with important implications in binding of the proteasome to ubiquitinylated substrates and subsequent protein degradation. In that regard, we are the first to provide biochemical evidence indicating that the R3IM motif of HsDSS1, in conjunction with 19S RP/20S CP (core particle) complex, regulates proteasome assembly and function through interaction with Rpn3/S3, and utilizes a specific subset of polyubiquitinated p53 as a substrate.